CA1120837A - Device for microbiological purposes - Google Patents
Device for microbiological purposesInfo
- Publication number
- CA1120837A CA1120837A CA000328718A CA328718A CA1120837A CA 1120837 A CA1120837 A CA 1120837A CA 000328718 A CA000328718 A CA 000328718A CA 328718 A CA328718 A CA 328718A CA 1120837 A CA1120837 A CA 1120837A
- Authority
- CA
- Canada
- Prior art keywords
- nutrient
- micro
- film
- card
- synthetic resin
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
- C12M41/30—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration
- C12M41/36—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration of biomass, e.g. colony counters or by turbidity measurements
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M23/00—Constructional details, e.g. recesses, hinges
- C12M23/02—Form or structure of the vessel
- C12M23/14—Bags
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S435/00—Chemistry: molecular biology and microbiology
- Y10S435/805—Test papers
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S435/00—Chemistry: molecular biology and microbiology
- Y10S435/81—Packaged device or kit
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- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Wood Science & Technology (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Biomedical Technology (AREA)
- Sustainable Development (AREA)
- Microbiology (AREA)
- Biotechnology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Clinical Laboratory Science (AREA)
- Analytical Chemistry (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
Abstract
ABSTRACT OF THE DISCLOSURE
A test unit device for microbiological investigation comprises a nutrient card and a covering layer thereover comprising a film of a water-insoluble synthetic resin modified with an opener. The film is formed from an aqueous dispersion of the resin and is impermeable to microorganisms but permeable to nutrients in the nutrient card, the device permits the ready determination of microorganism counts and the differentiation of microorganisms in diagnosis, it may also be employed in the production of an antibiogram and in the determination of the inhibiting concentration of antibiotics.
A test unit device for microbiological investigation comprises a nutrient card and a covering layer thereover comprising a film of a water-insoluble synthetic resin modified with an opener. The film is formed from an aqueous dispersion of the resin and is impermeable to microorganisms but permeable to nutrients in the nutrient card, the device permits the ready determination of microorganism counts and the differentiation of microorganisms in diagnosis, it may also be employed in the production of an antibiogram and in the determination of the inhibiting concentration of antibiotics.
Description
112V~37 The present invention is concerned with a test device for microbiological purposes; more especially the invention is concerned with such deviceq comprising a nutrient card.
The inveqtigation of certain body ~luids for the presence of micro-organisms, such aQ the detection of bacteria and fungi in urine, provides the physician, micro~iologist and the like with valuable indication~ for the diagnosis of particular diseases. Consequently, numerous test means have been developed in recent years for the detection of bacteria and fungi.
However, detection means of this type are also used fox the investigation o pharmaceuticals, focdstuffs, water, swimmung pools and the like where certain kinds of micro-organisms are harmful or can constitute a danger to health.
The known processes of culturing and detecting micro-organisms depend upon the fact that various selected nutrients and possibly selectively inhibiting agents are worked up with agar-agar to give a nutrient medium. Although nutrient media are important for special problems, they are disadvantageous for routine operation~, especially when cost-saving and comparatively large numbers of samples are to be investigated. The nutrient media must be prepared by the user in a relatively laborious manner from a dry powder.
Nutrient media ready for use are also commercially available in swollen form. However, nutrient agar media which are ready for use readily dry out and can then no longer be reversibly moistened, whîch means that they must be packed in moisture-proof packings which, in addition, take up a large volume.
.
d~
112V~37 More recently, especially for the investigation of foodstuffs and water, microbiological detection processes have been described in which nutrient cards are covered with membrane filters. For this purpose, the liquid to be investigated is filtered through the membrane filter, the membrane filter is laid upon a nutrient card moistened with sterile water or upon a filter card impregnated with sterile nutrient solution and this unit then cultured in a Petri dish. The micro-organismSretained on the membrane filter grow during the incubation period and become visible as colonies.
This process is only practicable when micro-organisms are to be detected in very small concentrations and, therefore, must be enriched before detection by means of incubation. The membrane filters and nutrient cards are provided loose and, because of the brittleness of the membrane material, require especially careful handling.
An improvement is provided by combining the membrane filter and the nutrient card into a single unit in a special frame (see German Patent 2,115,674, Charles P. Shaufus et al, published December 2, 1971).
The process according to the German Patent admittedly simplifie3 handling but the production of the unit comprising the membrane filter and the nutrient card in a special frame is laborious and expensive.
The dè~cribed filter processes are not suitable for medical diagnoqtic problems since a concentration of the micro-organisms on the s,urface of the filter is not necessary and, indeed, because of the necessity of distinguishing high micro-organism counts, preferably in the range of from 10 to 10 micro-organisms per ml. of body fluid, is of great dis-advantage.
1~2U83~
Furthermore, processes are known in which a micro-porous membrane is produced on a nutrient card in such a manner that the membrane material partly penetrates into the card (see German Patent ~o. 2,320,946, Billy Harper Haden et al, November 28, 1974~.
The process according to the German Offenlegungsschrift suffers from the disadvantage that the surface of the mem-brane has the same roughness or unevenness as that of the nutrient card. In the case of such a test, because of the unevenness of the surface, the micro-organisms are, without additional means, not visible at all or very poorly visible after the incubation. They must be rendered visible by colouring with dyestuffs. Therefore, the micro-organisms appear in a completely different manner to the viewer than in the case of the well-known detection processes on agar-agar. A visual differentiation of the species is not possible.
The colonies, which penetrate more or less into the nutrient card, cannot be separately used for inoculation. However, such an inoculation is absolutely essential for a differentiation of micro-organisms and for an investigation of resistance behaviour.
Another proces~ i9 known which is essentially a further development of the above-described process, it having the object of overcoming the disadvantages of this process.
This test, which is described in German Patent ~o. 2,320,943, Billy Harper Haden et al, September 16, 1976) is improved in that, instead of the membrane layer or above the membrane layer, there is applied a gel layer, the gel layer and the nutrient card passing over into one another.
It is thereby preferable also to incorporate nutrie~ts into the gel layer. As is known from experience, such gel layers cannot be ap~lied to paper surfaces with the same uniformity and smoothness as is possible in the case of swollen agar layers.
Therefore, this process ~uffers from disadvantages which are simllar to those possessed by the process according to German Patent No. 2,320,946.
Therefore, it is an object of the present invention to provide a dry nutrient substrate for micro-organisms which, upon dipping into a micro-organism-containing liquid to be investigated, takes up water with appropriate rapidity and the surface of which, in a moist state, is as smooth and uniform as that of the well-known agar media. The micro-organisms must remain on the surface, in the case of dipping into a micro~organism-containing solution, in an amount e~uiva-lent to their concentration. The micro-orgarisms must, within the usual incubation period (12 to 24 hours), form colonies as is the case with the well-known agar nutrient media. Thus, with these dry nutrient substrates, the number of micro-organisms and the species thereof must be capable of deter-mination in the ~ame manner as in the case of the well-known processes. Furthermore, it must be possible to test the resistance behaviour of the micro-organisms.
For solving this problem it was necessaxy to develop speci~l covering layers for covering the nutrient card which did not exhibit the disadvantages of the above-described proces~es.
It ha~, surprisingly, been found that films based upon synthetic resin dispersion~ provide a satisfactory solution to the problem. For this purpose, synthetic resin films are provided with "inner openers" which impart the following properties to films produced from the dispersions:
the film remains impermeable to micro-organisms, the film is permeable to micro-organi~m nutrients and to ilZ~)1!337 substances which selectively or generally inhibit or suppress the growth of micro-organisms the surface of the film i5 smooth;
on the surface of the film, the micro-organisms are firmly held corresponding to their concentration in the medium to be investigated.
In particular the invention is concerned with films in which the micro-organi~ms referred to in defining the properties of the film are bacteria.
Thus, according to the present invention, there is provided a device for microbiological purposes, comprising a nutrient card and a covering layer comprising a water-insoluble synthetic resin film modified with an opener, said opener being effective to render said film impermeable to micro-organisms but permeable to nutrients, said film having been produced from an aqueous dispersion of the synthetic resin.
In other aspects of the invention there is provided a method for determining micro-organism counts and for the differentiation of microorganisms; a method of producing an antibiogram, and a method for the determination of the minimum inhibiting concentration of antibiotics, all of which methods em?loy devices in accordance with the invention.
As the dispersed synthetic resin, there can, in principle, be used all water insoluble, synthetic resins dispersible in water, provided that the films formed from aqueous dispersions thereof do not have a negative effect on bacterial growth. Thus, for example, the following synthetic resin polymers can be employed: homopolymers of vinyl acetate, acrylic acid esters and the like; co-polymers of vinyl propionate and vinyl acetate, vinyl propionate and vinyl chloride, vinyl acetate and maleic acid esters, acrylic acid ~12V1~3~7 esters with acrylonitrile and vinyl propionate, hutadiene and styrene and the like.
As openers, there can, in principle, be used all water-soluble or water-swellable compounds which do not have a negative effect on bacterial growth, which dissolve or swell in the moistened film only to such an extent that the structure of the film is not so destroyed that the fllm breaks down or such large holes are formed that micro-organisms can pa~s through. If they are metabolised, then such meta-bolism must not provide the micro-organisms with the possibility of being able to pass through the film or of destroying the structure thereof to such an extent that the colonies are no longer clearly visible.
Of course, mixtures of substances which have opener properties can also be used. In special cases, the use of several openers in a film can be advantageous.
As openers there can preferably be used the follow-ing macromolecular substances: polyethylene glycols, poly-ethylene oxideY, polyvinylpyrrolidone, polyvinyl alcohols, partially saponified polyvinyl esters, copolymers of vinyl-pyrrolidone and vinyl esters, cellulose derivative3, for example, hydroxyalkylcelluloses, and the like. Furthermore, there can also be used low molecular weight materials, such as sugars, ~utrient salts and the like, as well as mixtures of such macro- and micro-molecular materials.
In the compo~ition of the formulation of the crude film mass, the addition of wetting agents is of advantage.
In this way, not only are the properties of the film improved but also the growth of certain species is selectively influenced and an undesired behaviour of the micro-organism during the incubation phase, for example, clustering of Protéus, is prevented. As wetting agents, there can be used 1120~33~
all substances conventionally employed in microbiology, for ex~nple, sodium dodecyl sulphate, sodium heptadecyl sulphate, N-cetyl-N,N,N-trimethylammonium bromide, polyoxyethylene ~orbitan monooleate and the like.
Since the growth of the micro-organisms is also decisively dependent upon the pH value of the medium on or in which they grow, with the help of conventional buffers, for example, Sorensen buffer, the pH value of the film mass is adjusted to the desired range. The salt concentration is thereby selected in such a manner that the osmolar conditions correspond to the requirements for a satisfactory micro-organism growth.
In addition to the openers, for example, macro-molecular compounds, wetting agents and buffers, the films can also have incorporated therein selective inhibiting materials or antibiotics so that, by means of such a formulation, the growth of micro-organisms can be selectively inhibited or the minimum inhibiting concentration of antibiotics can be determined.
Film3 of this type can be produced as follows:
The aqueou~ synthetic resin polymer dispersion i9 homogeneouQly stirred with an aqueous solution of the opener.
Into this aqueous mixture are stirred the other components, for example, the wetting agent, buffer, inhibiting materials and the like. In these crude film masses, the weight amount ratio of the dispersion polymer to the opener suitably varies from 50:1 to 1:5 and preferably 1:1 to 10:1. The addition of solvents depends upon the desired viscosity of the mass which, depending upon the method to be used for further working up, can vary very greatly. The concentrations of the remaining additives depend upon the requirements which they are to satisfy and i~2~)837 they can be present in amounts from 0 to 15%, by weight, and preferably from 1 to 5%, by weight, of the synthetic resin co~ering layer.
The films themselves are produced in the usual manner in that the crude film mass is coated, scraped, sprayed or the like, in a layer thickness of from 10 to lOOOu and preferably 50 to 200~, on to a substrate with which the crude film mass does not bind too firmly. After or during drying, the film is pulled or lifted off from the substrate and subsequently further worked up.
In order to stabilise films against stretching in the longitudinal or transverse direction, they can be produced by applying the crude film mass in known manner on to an inner carrier, for example, a woven or knitted material or the like, which lies upon an inert substrate. Preferred carriers are commercially available synthetic resin meshes of polyamide, polyester, polyethylene or the like which are known, for example, as bolting cloth and have, for example, a thickness of about 10 to 20ql.~ The inner carrier should, however, have a thickne~ less than the thickness of the film and be contained entirely within the film so as to ensure a smooth film outer surface.
Such ~uppoxted films can also be produced by impregnating or coating the inner carrier with the film mass, whereafter, by wiping off or the like, the film thick-ne~s can be adjusted. If such films are combined with known nutrient cards to give test systems, then a test device is obtained which fully satisfies the above-mentioned requirements.
~ utrient cards can be produced in known manner by impregnating filter papers or other appropriate absorbent carrier layers with solutions of nutrient mixtures which are conventional in microbiology and subsequently drying them.
~- 8 -il20~37 In this manner, a large variety of nutrient caxds can be pro-duced with various properties. Examples of nutrient media which can be used include MacConkey's nutrient medium, CLED
nutrient medium, Slanetz and Bartley'~ nutrient medium, China blue-lacto~e nutrient medium, endo nutrient medium, Wilson-Blair's bismuth sulphite nutrient medium, cetrimide nutrient medium, CaSo nutrient medium, Muller-Hintson's nutrient medium, Sabouraud's nutrient medium and the like.
The test device ready for use can be constructed in various ways. The model~ which have proved to be the best will now be described.
A self-supporting film or a film which is stabilised by an inner carrier iB laid upon a nutrient card. For the detection of micro-organisms, the nutrient card and the film are dipped into the micro-organism-containing solution to be investigated. Both are placed, one on top of the other, in a Petri dish and incubated for 12 to 24 hours. The micro-organisms to be detected become visible in the form of colonies in known manner~
If it is desired to quantify the number of micro-organisms, then it is preferable to produce the film in such a manner that, as inner carrier, a woven mesh is used in which, at definite di~tances, for example, of 0.5 cm. in each direction, the filaments are coloured in a different manner, the mesh thus being divided up into squares in a manner reminiscent of the engraved chamber of a haemocytometer. If such a film, together with the nutrient card, is dipped into a micro-organism-containing liquid and subsequently incubated, then, from the number of colonies per unit surface area, the number of micro-organisms per unit volume can be determined very easily. In the case of one device which is particularly easy to produce, the nutrient caxd is laid upon a stiff foil, ll~V~3~
a sornewhat wider piece of the above-described test film is laid thereover and the projecting part of the film is welded or stuck on to the foil in such a manner that the nutrient card is firmly clamped in the pocket resulting between the foil and the film. This test strip, which preferably has a width of 0.5 to 5 cm., in which the test area is preferably quadratic, is prepared for the user as follows: it is sealed or stuck between foils or coated papers and sterilised in conventional manner~ There is thu~ obtained a dry nutrient substrate in an extremely space-saving form. Furthermore, the nutrient substrate can be stored for a much longer period of time than a nutrient agar ~ince it cannot dry out and the sensitive nutrient materials in the dry state are much more stable than in swollen agar. Before use, the test strip is removed from the foil packin~, For the detection of micro-organism~, for example, in urine, the test strip is dipped in fresh, cleanly collected middle stream urine, i.e., urine being excreted before the bladder is empty, placed in an incubation vessel and incubated. The packing can possibly also serve as an incubation vessel. On the basis of comparative tables, the number of micro-organisms per ml. of urine can be deter-mined. It is preferable to combine a universal nutrient with ~everal selective nutrient media and/or selective nutrient media in one test lunit, For the preparatiqn of an antibiogram, such a device which contains, for example, a universal nutrient card, for example, Muller-Hinton nutrient medium, is moistened with sterile water, a slurry of the micro-organisms to ~e tested is applied thereto with a spatula and an antibiotic leaflet, such as is described in German Industrial Standard No. S8940, iR placed thereon. After in!cubation, evaluation is carried out in the usual way.
112~837 Another possibility of inve~tigating the resistance behaviour of micro-organism~ to an antibiotic is the deter-mination of the minimum inhibiting concentration (MIC). This i9 determined in the following manner:
The antibiotic in the desired concentration is incorporated directly into the film mass and test strips ara produced from the films prepared therefrom. A nutrient card with a universal nutrient medium is, together therewith, placed on to a foil and the appropriate antibiotic-containing film sealed thereover in the manner described above. It is also possible to incorporate the antibiotic into the nutrient card.
Some antibiotics are only stable for a comparatively long period of time in a relatively narrow p~ range, this pH
range being one which doe~ not necessarily correspond to the optimum pH for the growth of the micro-organisms. For the determination of the MIC of such substances by the process according to the pre~ent invention, the films are produced in the following manner and further worked up to give test ~trips in the manner described above.
On to one of the above-described antibiotic-free films there is applied a second film which contains the desired antibiotic. ~he buffer capacity of the second film is adjusted in such a manner that, upon dipping into the aqueous sample material to be investigated, the buffer of the first film or of the nutrient medium ensures the adjustment of the p~ value to one which is optimum for the growth of the micro-organism. The second film can be produced from the same crude film mass a~ the first film, except that it must have a different pH value corresponding to that desired for the antibiotic. However, it is not necessary that the synthetic resin dispersion and the opener are identical in both films.
ilZ~837 It iR also po~sible to produce the second film, in which the antibiotic is incorporated, from a water-soluble or water-swellable film former which is substantially inert for the growth of the micro-organism~. As film formers for this purpose, there are preferably used the same materials as can also be used as openers.
Furthermore, the antibiotic-containing film and the nutrient card can, of course, also have different pH values so that the buffer of the nutrient card ensures the optimum pH value for the growth of the micro-organism~ only after moistening the surface of the film.
Incorporating the antibiotic directly into the film or nutrient card and incorporating it into the test unit for the determination of the MIC has the following advantages: It is no longer necessary to lay antibiotic leaflets on to an agar plate. Without the previous unavoid-able intermediate step of micro-organism culturing, the resistance behaviour of the micro-organisms can be tested directly from the sample material. This novel method avoids the complicated process, previously necessary for the deter-mination of the MIC, of having to weigh out the antibiotic every time for each series of investigations. Finally, the assessment of the inhibited growth of the micro-organisms is substantially easier: it 19 now no longer necessary to assess the degree of turbidity of the nutrient broth: on the co~trary, the growth of the micro-organisms can be evaluated directly on the surface of the film and classified thereon.
The in~ention is illustrated in a particular embodiment by reference to the accompanying drawings in which:
33~
FIGURE 1 show~ a front view of a test strip according to the invention, and FIGURE 2 is a side view of an individually sealed test device of the invention.
With further reference to Figure 1, a test strip 10 comprise~ a pair of spaced apart nutrient cards 1, a covering layer 2 lying thereover and a foil 3. The nutrient cardsl and the overlying portion~ of covering layer 2 define test zones 5 and 6.
The nutrient cards 1 are dispo~ed between covering layer 2 and foil 3, layer 2 being adhered to foil 3 in zones 7, 8 and 9 so as to firmly locate the cards 1.
Foil 3 includes a handle portion 12.
With further reference to Figure 2, a test device 20 comprises four nutrient cards 21, a pair of covering layers 22 and a pair of foils 23. The device 20 is sealed in a packing 24.
The nutrient cards 21 and the overlying portions of covering layers 22 define four test zones 25, 26, 27 and 28.
The foi~ 23 are adhered together and the layers 22 are adhered to the foils 23 in zones 29, 30, 31, 32, 33 and .
34, peripheral to cards 21, so as to firmly locate cards 21.
The foil 23 includes a handle portion 35.
,~ , The following Exampqes are given for the purpose of illustrating the present invention in particular and preferred embodiments.
Example 1 Production of films A large variety of differ~nt synthetic resin dis~
persions and openers can be used for the production of the films. The two components are, together with adjuvants, stirred to give a crude film mass. (The openers are water-soluble, 112VB3~
relatively slightly ~welling, high polymer substances).
There is demonstrated the multiplicity of com-bination possibilities of the raw materials which permit the production of u~eful films.
The openers are dissolved, with stirring, in water to which is added the synthetic resin dispersions in the amounts ~tated in the following Tables, this mixture being stirred until homogeneous. ~hereafter, 1 g. of wetting agent is added thereto, the choice of wetting agent depending upon the micro-organi~ms which are to be detected, for example, sodium dodecyl sulphate for the inhibition of gram-negative flora, polyoxyethylene~orbitan oleate as additive for the detection of Lactobacillae and for inhibiting the clumping or agglomeration of micro-organism~, sodium heptadecyl sulphate for the inhibition of gram-po~itive flora and in media for the enrichment of coliform micro-organisms and N-cetyl-~,~,N-trimethylammonium bromide for the selective enrichment of Pseudomonas. In addition, 1 ml. of a buffered sodium chloride solution (composition: 92 g~ disodium hydrogen phosphate dihydrate,2Ll g. potassium dihydrogen phosphate and 7.65 g. sodium chloride in 1000 ml. water) is added thereto and the p~ adjusted to 6.9 to 7.5 with 0.1 aqueou~ ~odium hydroxide ~olution.
- 14 _ 1~2~ 7 Tabulation of the amountq of dispersion~ opener and water . I
Dis~ersion:
Homopolymer of acrylic acid e~ter 60 g, 50 g, 50 g.
O~ener:
Hydroxypropylmethylcellulose 4 g. _ -Copolymers of ethylene glycol and ethylene oxide _ 18 g, Polyvinylpyrrolidone _ _ 24 g.
Water 30 ml. 26 ml. 31 ml.
l~LZ0~33'7 o~
_ _ I
~ I I I I I In ~
o . ~
o I I I I I In o~
'~J ~ ~
o o~ U~ , , ~ . ~
, :~ . .
. ~ .
: :
~ , , , , , ~ ~
~, , ,, , _ _ ',~ ~
,~ ~.
' ~ o .~.~ o o ~ o . .~ ~o ~ C ~ o ,~ o ~ o ~ ~,., 3 ~ ~ . ~ ch 1~ ~ O O ~ O I D O G ~ o ~ ~ ~ l 112083~7 . . , Dispersion:
Copolymer of butadiene 52 g. 74 g. 62 g. 62 g.
and styrene Opener:
Polyvinylpyrrolidone 16 g, 16 g, ~
Copolymer of ethylene glycol and ethylene oxide _ _ 16 g, Copolymer of vinyl acetate and vinyl ~-pyrrolidone _ _ _ 16 g.
Water 22ml. 22ml. 22ml. 47ml.
Dispersion:
Homopolymer of vinyl 62 g, 62 g, acetate Opener- _ .
Polyvinylpyrxolidone 20 g.
Copolymer of vinyl .
acetate and vinyl pyrrolidone _ 16 g.
Water 26ml. 47ml.
l~ZI~&3~7 - ~ l - -Dispersion:
Copolymer of acrylonitrile, acrylic acid ester and vinyl 78 g. 92 g, 78 g.
propionate ~)ener:
Polyvinylpyrrolidone 16 g. ¦ 16 g.
Copolymer of ethylene li glycol and ethylene oxide _ _ 16 g.
Water 22ml. 1 22ml. 23ml.
.
Dispersion: .
Copolymer of maleic acid and vinyl acetate 58 g.60 g.60 g.
OPener:
Polyvinylpyrrolidone 13 g. _ Polyethylene oxide _ 16 g. _ Copolymer of vinyl acetate and vinyl pyrrolidone _ _ 16 g.
Water 18ml.62ml.47ml. , llZV~37 Dispersion:
~Iomopolymer of vin~l chloride 62 g .1 Opener:
Polyethylene oxide 16 g.
Water 62ml.
_ Dispersion:
Copolymer of vinyl chloride and vinyl propionate 52 g, Opener:
Polyvinylpyrrolidone 15.7 g Water 22ml.
The crude film mass is scraped on to an inert substrate in known manner with a layer thickness of 30ql and dried at a temperature of 50 to 70C. After drying, the film is cut up into the desired size and further worked up.
Example 2 Production of a test strip for the determination of the micro-orqanism count in urine and for the rouqh differentiation of micro-orqan_sms.
62 g. of a dispersion of a co-polymer of vinyl acetate and vinyl propionate are stirred with a solution of 16 g. polyvinylpyrrolidone in 22 ml. water. Then, as described in Example 1, 1 ml. of a buffered physiological sodium chloride solution and 1 ml. polyoxyethylene sorbitan monooleate are stirred therein and the pH adjusted to 7.3 with aqueous sodium hydroxide solution.
A nylon mesh with a thickness of 5ql is laid on to a polyethylene foil, upon which the film mass is scraped with a layer thickness of 35ql. After drying at 60C., the support film now obtained is stripped off from the foil and cut up into 6 cm. wide bands.
llZ~837 On to a 2001 thick foil with the dimensions of
The inveqtigation of certain body ~luids for the presence of micro-organisms, such aQ the detection of bacteria and fungi in urine, provides the physician, micro~iologist and the like with valuable indication~ for the diagnosis of particular diseases. Consequently, numerous test means have been developed in recent years for the detection of bacteria and fungi.
However, detection means of this type are also used fox the investigation o pharmaceuticals, focdstuffs, water, swimmung pools and the like where certain kinds of micro-organisms are harmful or can constitute a danger to health.
The known processes of culturing and detecting micro-organisms depend upon the fact that various selected nutrients and possibly selectively inhibiting agents are worked up with agar-agar to give a nutrient medium. Although nutrient media are important for special problems, they are disadvantageous for routine operation~, especially when cost-saving and comparatively large numbers of samples are to be investigated. The nutrient media must be prepared by the user in a relatively laborious manner from a dry powder.
Nutrient media ready for use are also commercially available in swollen form. However, nutrient agar media which are ready for use readily dry out and can then no longer be reversibly moistened, whîch means that they must be packed in moisture-proof packings which, in addition, take up a large volume.
.
d~
112V~37 More recently, especially for the investigation of foodstuffs and water, microbiological detection processes have been described in which nutrient cards are covered with membrane filters. For this purpose, the liquid to be investigated is filtered through the membrane filter, the membrane filter is laid upon a nutrient card moistened with sterile water or upon a filter card impregnated with sterile nutrient solution and this unit then cultured in a Petri dish. The micro-organismSretained on the membrane filter grow during the incubation period and become visible as colonies.
This process is only practicable when micro-organisms are to be detected in very small concentrations and, therefore, must be enriched before detection by means of incubation. The membrane filters and nutrient cards are provided loose and, because of the brittleness of the membrane material, require especially careful handling.
An improvement is provided by combining the membrane filter and the nutrient card into a single unit in a special frame (see German Patent 2,115,674, Charles P. Shaufus et al, published December 2, 1971).
The process according to the German Patent admittedly simplifie3 handling but the production of the unit comprising the membrane filter and the nutrient card in a special frame is laborious and expensive.
The dè~cribed filter processes are not suitable for medical diagnoqtic problems since a concentration of the micro-organisms on the s,urface of the filter is not necessary and, indeed, because of the necessity of distinguishing high micro-organism counts, preferably in the range of from 10 to 10 micro-organisms per ml. of body fluid, is of great dis-advantage.
1~2U83~
Furthermore, processes are known in which a micro-porous membrane is produced on a nutrient card in such a manner that the membrane material partly penetrates into the card (see German Patent ~o. 2,320,946, Billy Harper Haden et al, November 28, 1974~.
The process according to the German Offenlegungsschrift suffers from the disadvantage that the surface of the mem-brane has the same roughness or unevenness as that of the nutrient card. In the case of such a test, because of the unevenness of the surface, the micro-organisms are, without additional means, not visible at all or very poorly visible after the incubation. They must be rendered visible by colouring with dyestuffs. Therefore, the micro-organisms appear in a completely different manner to the viewer than in the case of the well-known detection processes on agar-agar. A visual differentiation of the species is not possible.
The colonies, which penetrate more or less into the nutrient card, cannot be separately used for inoculation. However, such an inoculation is absolutely essential for a differentiation of micro-organisms and for an investigation of resistance behaviour.
Another proces~ i9 known which is essentially a further development of the above-described process, it having the object of overcoming the disadvantages of this process.
This test, which is described in German Patent ~o. 2,320,943, Billy Harper Haden et al, September 16, 1976) is improved in that, instead of the membrane layer or above the membrane layer, there is applied a gel layer, the gel layer and the nutrient card passing over into one another.
It is thereby preferable also to incorporate nutrie~ts into the gel layer. As is known from experience, such gel layers cannot be ap~lied to paper surfaces with the same uniformity and smoothness as is possible in the case of swollen agar layers.
Therefore, this process ~uffers from disadvantages which are simllar to those possessed by the process according to German Patent No. 2,320,946.
Therefore, it is an object of the present invention to provide a dry nutrient substrate for micro-organisms which, upon dipping into a micro-organism-containing liquid to be investigated, takes up water with appropriate rapidity and the surface of which, in a moist state, is as smooth and uniform as that of the well-known agar media. The micro-organisms must remain on the surface, in the case of dipping into a micro~organism-containing solution, in an amount e~uiva-lent to their concentration. The micro-orgarisms must, within the usual incubation period (12 to 24 hours), form colonies as is the case with the well-known agar nutrient media. Thus, with these dry nutrient substrates, the number of micro-organisms and the species thereof must be capable of deter-mination in the ~ame manner as in the case of the well-known processes. Furthermore, it must be possible to test the resistance behaviour of the micro-organisms.
For solving this problem it was necessaxy to develop speci~l covering layers for covering the nutrient card which did not exhibit the disadvantages of the above-described proces~es.
It ha~, surprisingly, been found that films based upon synthetic resin dispersion~ provide a satisfactory solution to the problem. For this purpose, synthetic resin films are provided with "inner openers" which impart the following properties to films produced from the dispersions:
the film remains impermeable to micro-organisms, the film is permeable to micro-organi~m nutrients and to ilZ~)1!337 substances which selectively or generally inhibit or suppress the growth of micro-organisms the surface of the film i5 smooth;
on the surface of the film, the micro-organisms are firmly held corresponding to their concentration in the medium to be investigated.
In particular the invention is concerned with films in which the micro-organi~ms referred to in defining the properties of the film are bacteria.
Thus, according to the present invention, there is provided a device for microbiological purposes, comprising a nutrient card and a covering layer comprising a water-insoluble synthetic resin film modified with an opener, said opener being effective to render said film impermeable to micro-organisms but permeable to nutrients, said film having been produced from an aqueous dispersion of the synthetic resin.
In other aspects of the invention there is provided a method for determining micro-organism counts and for the differentiation of microorganisms; a method of producing an antibiogram, and a method for the determination of the minimum inhibiting concentration of antibiotics, all of which methods em?loy devices in accordance with the invention.
As the dispersed synthetic resin, there can, in principle, be used all water insoluble, synthetic resins dispersible in water, provided that the films formed from aqueous dispersions thereof do not have a negative effect on bacterial growth. Thus, for example, the following synthetic resin polymers can be employed: homopolymers of vinyl acetate, acrylic acid esters and the like; co-polymers of vinyl propionate and vinyl acetate, vinyl propionate and vinyl chloride, vinyl acetate and maleic acid esters, acrylic acid ~12V1~3~7 esters with acrylonitrile and vinyl propionate, hutadiene and styrene and the like.
As openers, there can, in principle, be used all water-soluble or water-swellable compounds which do not have a negative effect on bacterial growth, which dissolve or swell in the moistened film only to such an extent that the structure of the film is not so destroyed that the fllm breaks down or such large holes are formed that micro-organisms can pa~s through. If they are metabolised, then such meta-bolism must not provide the micro-organisms with the possibility of being able to pass through the film or of destroying the structure thereof to such an extent that the colonies are no longer clearly visible.
Of course, mixtures of substances which have opener properties can also be used. In special cases, the use of several openers in a film can be advantageous.
As openers there can preferably be used the follow-ing macromolecular substances: polyethylene glycols, poly-ethylene oxideY, polyvinylpyrrolidone, polyvinyl alcohols, partially saponified polyvinyl esters, copolymers of vinyl-pyrrolidone and vinyl esters, cellulose derivative3, for example, hydroxyalkylcelluloses, and the like. Furthermore, there can also be used low molecular weight materials, such as sugars, ~utrient salts and the like, as well as mixtures of such macro- and micro-molecular materials.
In the compo~ition of the formulation of the crude film mass, the addition of wetting agents is of advantage.
In this way, not only are the properties of the film improved but also the growth of certain species is selectively influenced and an undesired behaviour of the micro-organism during the incubation phase, for example, clustering of Protéus, is prevented. As wetting agents, there can be used 1120~33~
all substances conventionally employed in microbiology, for ex~nple, sodium dodecyl sulphate, sodium heptadecyl sulphate, N-cetyl-N,N,N-trimethylammonium bromide, polyoxyethylene ~orbitan monooleate and the like.
Since the growth of the micro-organisms is also decisively dependent upon the pH value of the medium on or in which they grow, with the help of conventional buffers, for example, Sorensen buffer, the pH value of the film mass is adjusted to the desired range. The salt concentration is thereby selected in such a manner that the osmolar conditions correspond to the requirements for a satisfactory micro-organism growth.
In addition to the openers, for example, macro-molecular compounds, wetting agents and buffers, the films can also have incorporated therein selective inhibiting materials or antibiotics so that, by means of such a formulation, the growth of micro-organisms can be selectively inhibited or the minimum inhibiting concentration of antibiotics can be determined.
Film3 of this type can be produced as follows:
The aqueou~ synthetic resin polymer dispersion i9 homogeneouQly stirred with an aqueous solution of the opener.
Into this aqueous mixture are stirred the other components, for example, the wetting agent, buffer, inhibiting materials and the like. In these crude film masses, the weight amount ratio of the dispersion polymer to the opener suitably varies from 50:1 to 1:5 and preferably 1:1 to 10:1. The addition of solvents depends upon the desired viscosity of the mass which, depending upon the method to be used for further working up, can vary very greatly. The concentrations of the remaining additives depend upon the requirements which they are to satisfy and i~2~)837 they can be present in amounts from 0 to 15%, by weight, and preferably from 1 to 5%, by weight, of the synthetic resin co~ering layer.
The films themselves are produced in the usual manner in that the crude film mass is coated, scraped, sprayed or the like, in a layer thickness of from 10 to lOOOu and preferably 50 to 200~, on to a substrate with which the crude film mass does not bind too firmly. After or during drying, the film is pulled or lifted off from the substrate and subsequently further worked up.
In order to stabilise films against stretching in the longitudinal or transverse direction, they can be produced by applying the crude film mass in known manner on to an inner carrier, for example, a woven or knitted material or the like, which lies upon an inert substrate. Preferred carriers are commercially available synthetic resin meshes of polyamide, polyester, polyethylene or the like which are known, for example, as bolting cloth and have, for example, a thickness of about 10 to 20ql.~ The inner carrier should, however, have a thickne~ less than the thickness of the film and be contained entirely within the film so as to ensure a smooth film outer surface.
Such ~uppoxted films can also be produced by impregnating or coating the inner carrier with the film mass, whereafter, by wiping off or the like, the film thick-ne~s can be adjusted. If such films are combined with known nutrient cards to give test systems, then a test device is obtained which fully satisfies the above-mentioned requirements.
~ utrient cards can be produced in known manner by impregnating filter papers or other appropriate absorbent carrier layers with solutions of nutrient mixtures which are conventional in microbiology and subsequently drying them.
~- 8 -il20~37 In this manner, a large variety of nutrient caxds can be pro-duced with various properties. Examples of nutrient media which can be used include MacConkey's nutrient medium, CLED
nutrient medium, Slanetz and Bartley'~ nutrient medium, China blue-lacto~e nutrient medium, endo nutrient medium, Wilson-Blair's bismuth sulphite nutrient medium, cetrimide nutrient medium, CaSo nutrient medium, Muller-Hintson's nutrient medium, Sabouraud's nutrient medium and the like.
The test device ready for use can be constructed in various ways. The model~ which have proved to be the best will now be described.
A self-supporting film or a film which is stabilised by an inner carrier iB laid upon a nutrient card. For the detection of micro-organisms, the nutrient card and the film are dipped into the micro-organism-containing solution to be investigated. Both are placed, one on top of the other, in a Petri dish and incubated for 12 to 24 hours. The micro-organisms to be detected become visible in the form of colonies in known manner~
If it is desired to quantify the number of micro-organisms, then it is preferable to produce the film in such a manner that, as inner carrier, a woven mesh is used in which, at definite di~tances, for example, of 0.5 cm. in each direction, the filaments are coloured in a different manner, the mesh thus being divided up into squares in a manner reminiscent of the engraved chamber of a haemocytometer. If such a film, together with the nutrient card, is dipped into a micro-organism-containing liquid and subsequently incubated, then, from the number of colonies per unit surface area, the number of micro-organisms per unit volume can be determined very easily. In the case of one device which is particularly easy to produce, the nutrient caxd is laid upon a stiff foil, ll~V~3~
a sornewhat wider piece of the above-described test film is laid thereover and the projecting part of the film is welded or stuck on to the foil in such a manner that the nutrient card is firmly clamped in the pocket resulting between the foil and the film. This test strip, which preferably has a width of 0.5 to 5 cm., in which the test area is preferably quadratic, is prepared for the user as follows: it is sealed or stuck between foils or coated papers and sterilised in conventional manner~ There is thu~ obtained a dry nutrient substrate in an extremely space-saving form. Furthermore, the nutrient substrate can be stored for a much longer period of time than a nutrient agar ~ince it cannot dry out and the sensitive nutrient materials in the dry state are much more stable than in swollen agar. Before use, the test strip is removed from the foil packin~, For the detection of micro-organism~, for example, in urine, the test strip is dipped in fresh, cleanly collected middle stream urine, i.e., urine being excreted before the bladder is empty, placed in an incubation vessel and incubated. The packing can possibly also serve as an incubation vessel. On the basis of comparative tables, the number of micro-organisms per ml. of urine can be deter-mined. It is preferable to combine a universal nutrient with ~everal selective nutrient media and/or selective nutrient media in one test lunit, For the preparatiqn of an antibiogram, such a device which contains, for example, a universal nutrient card, for example, Muller-Hinton nutrient medium, is moistened with sterile water, a slurry of the micro-organisms to ~e tested is applied thereto with a spatula and an antibiotic leaflet, such as is described in German Industrial Standard No. S8940, iR placed thereon. After in!cubation, evaluation is carried out in the usual way.
112~837 Another possibility of inve~tigating the resistance behaviour of micro-organism~ to an antibiotic is the deter-mination of the minimum inhibiting concentration (MIC). This i9 determined in the following manner:
The antibiotic in the desired concentration is incorporated directly into the film mass and test strips ara produced from the films prepared therefrom. A nutrient card with a universal nutrient medium is, together therewith, placed on to a foil and the appropriate antibiotic-containing film sealed thereover in the manner described above. It is also possible to incorporate the antibiotic into the nutrient card.
Some antibiotics are only stable for a comparatively long period of time in a relatively narrow p~ range, this pH
range being one which doe~ not necessarily correspond to the optimum pH for the growth of the micro-organisms. For the determination of the MIC of such substances by the process according to the pre~ent invention, the films are produced in the following manner and further worked up to give test ~trips in the manner described above.
On to one of the above-described antibiotic-free films there is applied a second film which contains the desired antibiotic. ~he buffer capacity of the second film is adjusted in such a manner that, upon dipping into the aqueous sample material to be investigated, the buffer of the first film or of the nutrient medium ensures the adjustment of the p~ value to one which is optimum for the growth of the micro-organism. The second film can be produced from the same crude film mass a~ the first film, except that it must have a different pH value corresponding to that desired for the antibiotic. However, it is not necessary that the synthetic resin dispersion and the opener are identical in both films.
ilZ~837 It iR also po~sible to produce the second film, in which the antibiotic is incorporated, from a water-soluble or water-swellable film former which is substantially inert for the growth of the micro-organism~. As film formers for this purpose, there are preferably used the same materials as can also be used as openers.
Furthermore, the antibiotic-containing film and the nutrient card can, of course, also have different pH values so that the buffer of the nutrient card ensures the optimum pH value for the growth of the micro-organism~ only after moistening the surface of the film.
Incorporating the antibiotic directly into the film or nutrient card and incorporating it into the test unit for the determination of the MIC has the following advantages: It is no longer necessary to lay antibiotic leaflets on to an agar plate. Without the previous unavoid-able intermediate step of micro-organism culturing, the resistance behaviour of the micro-organisms can be tested directly from the sample material. This novel method avoids the complicated process, previously necessary for the deter-mination of the MIC, of having to weigh out the antibiotic every time for each series of investigations. Finally, the assessment of the inhibited growth of the micro-organisms is substantially easier: it 19 now no longer necessary to assess the degree of turbidity of the nutrient broth: on the co~trary, the growth of the micro-organisms can be evaluated directly on the surface of the film and classified thereon.
The in~ention is illustrated in a particular embodiment by reference to the accompanying drawings in which:
33~
FIGURE 1 show~ a front view of a test strip according to the invention, and FIGURE 2 is a side view of an individually sealed test device of the invention.
With further reference to Figure 1, a test strip 10 comprise~ a pair of spaced apart nutrient cards 1, a covering layer 2 lying thereover and a foil 3. The nutrient cardsl and the overlying portion~ of covering layer 2 define test zones 5 and 6.
The nutrient cards 1 are dispo~ed between covering layer 2 and foil 3, layer 2 being adhered to foil 3 in zones 7, 8 and 9 so as to firmly locate the cards 1.
Foil 3 includes a handle portion 12.
With further reference to Figure 2, a test device 20 comprises four nutrient cards 21, a pair of covering layers 22 and a pair of foils 23. The device 20 is sealed in a packing 24.
The nutrient cards 21 and the overlying portions of covering layers 22 define four test zones 25, 26, 27 and 28.
The foi~ 23 are adhered together and the layers 22 are adhered to the foils 23 in zones 29, 30, 31, 32, 33 and .
34, peripheral to cards 21, so as to firmly locate cards 21.
The foil 23 includes a handle portion 35.
,~ , The following Exampqes are given for the purpose of illustrating the present invention in particular and preferred embodiments.
Example 1 Production of films A large variety of differ~nt synthetic resin dis~
persions and openers can be used for the production of the films. The two components are, together with adjuvants, stirred to give a crude film mass. (The openers are water-soluble, 112VB3~
relatively slightly ~welling, high polymer substances).
There is demonstrated the multiplicity of com-bination possibilities of the raw materials which permit the production of u~eful films.
The openers are dissolved, with stirring, in water to which is added the synthetic resin dispersions in the amounts ~tated in the following Tables, this mixture being stirred until homogeneous. ~hereafter, 1 g. of wetting agent is added thereto, the choice of wetting agent depending upon the micro-organi~ms which are to be detected, for example, sodium dodecyl sulphate for the inhibition of gram-negative flora, polyoxyethylene~orbitan oleate as additive for the detection of Lactobacillae and for inhibiting the clumping or agglomeration of micro-organism~, sodium heptadecyl sulphate for the inhibition of gram-po~itive flora and in media for the enrichment of coliform micro-organisms and N-cetyl-~,~,N-trimethylammonium bromide for the selective enrichment of Pseudomonas. In addition, 1 ml. of a buffered sodium chloride solution (composition: 92 g~ disodium hydrogen phosphate dihydrate,2Ll g. potassium dihydrogen phosphate and 7.65 g. sodium chloride in 1000 ml. water) is added thereto and the p~ adjusted to 6.9 to 7.5 with 0.1 aqueou~ ~odium hydroxide ~olution.
- 14 _ 1~2~ 7 Tabulation of the amountq of dispersion~ opener and water . I
Dis~ersion:
Homopolymer of acrylic acid e~ter 60 g, 50 g, 50 g.
O~ener:
Hydroxypropylmethylcellulose 4 g. _ -Copolymers of ethylene glycol and ethylene oxide _ 18 g, Polyvinylpyrrolidone _ _ 24 g.
Water 30 ml. 26 ml. 31 ml.
l~LZ0~33'7 o~
_ _ I
~ I I I I I In ~
o . ~
o I I I I I In o~
'~J ~ ~
o o~ U~ , , ~ . ~
, :~ . .
. ~ .
: :
~ , , , , , ~ ~
~, , ,, , _ _ ',~ ~
,~ ~.
' ~ o .~.~ o o ~ o . .~ ~o ~ C ~ o ,~ o ~ o ~ ~,., 3 ~ ~ . ~ ch 1~ ~ O O ~ O I D O G ~ o ~ ~ ~ l 112083~7 . . , Dispersion:
Copolymer of butadiene 52 g. 74 g. 62 g. 62 g.
and styrene Opener:
Polyvinylpyrrolidone 16 g, 16 g, ~
Copolymer of ethylene glycol and ethylene oxide _ _ 16 g, Copolymer of vinyl acetate and vinyl ~-pyrrolidone _ _ _ 16 g.
Water 22ml. 22ml. 22ml. 47ml.
Dispersion:
Homopolymer of vinyl 62 g, 62 g, acetate Opener- _ .
Polyvinylpyrxolidone 20 g.
Copolymer of vinyl .
acetate and vinyl pyrrolidone _ 16 g.
Water 26ml. 47ml.
l~ZI~&3~7 - ~ l - -Dispersion:
Copolymer of acrylonitrile, acrylic acid ester and vinyl 78 g. 92 g, 78 g.
propionate ~)ener:
Polyvinylpyrrolidone 16 g. ¦ 16 g.
Copolymer of ethylene li glycol and ethylene oxide _ _ 16 g.
Water 22ml. 1 22ml. 23ml.
.
Dispersion: .
Copolymer of maleic acid and vinyl acetate 58 g.60 g.60 g.
OPener:
Polyvinylpyrrolidone 13 g. _ Polyethylene oxide _ 16 g. _ Copolymer of vinyl acetate and vinyl pyrrolidone _ _ 16 g.
Water 18ml.62ml.47ml. , llZV~37 Dispersion:
~Iomopolymer of vin~l chloride 62 g .1 Opener:
Polyethylene oxide 16 g.
Water 62ml.
_ Dispersion:
Copolymer of vinyl chloride and vinyl propionate 52 g, Opener:
Polyvinylpyrrolidone 15.7 g Water 22ml.
The crude film mass is scraped on to an inert substrate in known manner with a layer thickness of 30ql and dried at a temperature of 50 to 70C. After drying, the film is cut up into the desired size and further worked up.
Example 2 Production of a test strip for the determination of the micro-orqanism count in urine and for the rouqh differentiation of micro-orqan_sms.
62 g. of a dispersion of a co-polymer of vinyl acetate and vinyl propionate are stirred with a solution of 16 g. polyvinylpyrrolidone in 22 ml. water. Then, as described in Example 1, 1 ml. of a buffered physiological sodium chloride solution and 1 ml. polyoxyethylene sorbitan monooleate are stirred therein and the pH adjusted to 7.3 with aqueous sodium hydroxide solution.
A nylon mesh with a thickness of 5ql is laid on to a polyethylene foil, upon which the film mass is scraped with a layer thickness of 35ql. After drying at 60C., the support film now obtained is stripped off from the foil and cut up into 6 cm. wide bands.
llZ~837 On to a 2001 thick foil with the dimensions of
2 x 12 cm. there are now successively laid two nutrient cards with the dimensions of 2 x 2 cm~ containing Ca-So medium and ~acConkey's medium. Over the two nutrient cards is laid the film and attached thereto by sticking on to the foil (see Figures 1 and 2 of the accompanying drawings). A second test strip is produced according to the process just described in which, as nutrient media, there are used nutrient cards with Pseudomonas elective arld Entrococci selective media -The two test strips are stuck together to give a test strip tsee Figures 1 and 2 of the accompanying drawings). In order to protect the test strip against contamination and moisture, it is individually sealed, for example, with polyethylene-coated paper, and sterilised in known manner (see Figure 2 of the accompanying drawings). For the detection of micro-organi ms in urine, the test strip is removed from the packing, dipped into fresh middle stream urine and incubated in an incubation vessel for 12 to 24 hours at 37C. The pathogenic micro-organisms present in the urine manifest, on such a test strip, the same growth behaviour as on the well-known agar nutrient media.
Exam~le 3 Determination and differentiation of micro-orqanisms Films produced in the manner described in Example 1 are cut up into squares with 5 cm. long sides.
~utrient cards of the same size are laid in a Petri dish and moistened with sterile water. As nutrient media, there can be used all nutrient and nutrient-inhibitor compositions known in microbiology, For the determinatlon of the micro-organism count, it is best to use a universal medium, for example, the Ca-So medium. For micro-organism differentiation, there is used an 1~2V83'7 elective or selective medium, for example MacConkey's medium for the substantial inhibition of the gram~positive flora, Slanetz and Bartley's enterococcal-selective medium, Pseudo-monas elective medium, Wilson and Blair's Salmonella selective medium and the like~
On to the moistened nutrient cards are laid films, the wetting agents of which are chosen in such a manner that they are compatible with the nutrient medium and a precisely pipetted amount of micro-organism suspension distributed thereon with a spatula.
After an incubation period of 12 to 48 hours at 30 to 37C., by counting the colonies, the micro-organism count can be determined and, by the growth behaviour of the micro-organisms on the various media, the type thereof can be identified.
Example 4 Diaqnosis of urinarY infections - For the diagnosis of a urinary infection, use is made of a test strip according to Example 1. The wetting agent of the film i9, as in Example 3, coordinated with the nutrient medium. For this purpose, a nutrient card is cut with the dimensions of 2.5 x 2.5 cm., laid on a 2.5 x 10 cm.
~ealing foil and the film sealed with a hot-seal roller in such a manner that it lies close to the nutrient card. It is preferable to employ one nutrient card with CLED medium and one with MacConkey medium, which are placed side by side.
When such a test strip is dipped into a freshly excreted middle stream of urine and then incubated, there are found for all pathogenic micro-org~nisms in the urine, especially Colibacteria, Enterococci, Proteus mirabilis, Proteus vulqaris, Proteus morqanii, Proteus rettqeri, Pseudomonas aeruqinosa, Candida albicans, Klebsiella, Aerobacter aeroqenes, 11.ZC~337 ~ ilococcus aureus and Citrobacter, the same results as -with the well-known nutrient agar. The great advantage of the new test system in comparison with agar is that the test strips are packed in a sterile manner, can be stored for year~ and only require a fraction of the storage space needed for agar tests and, in addition, destruction thereof by incineration is less laborious.
Example 5 Preparation of antibioqram~
In order to test the resistance behaviour of micro-organisms cultured according to Example 4, the following procedure is used:
The usual procedure i9 employed in which colonies are inoculated with a loop, slurried in sterîle physiological sodium chloride solution and 0.1 ml. thereof further worked up.
For this purpose, a round nutrient card, for example, contain-ing Muller-Hinton's medium, is moistened with sterila water, covered with a film of the same size, the suspension of micro-organism spread thereon with a spatula and an antibiotic test leaflet according to German Industrial Standard ~o.
58940 laid thereon. In contradistinction to nutrient agar, it is not necessary to press on the test leaflet since this itself adheres to the moist surface of the test strip. After in¢ubation, for all pathogenic micro-organisms of the urine there are observed, in the case of all antibiotics used, a resistance behaviour which is identical with incubation in normal agar nutrient substrates.
- 2~ -112~83~
Example_6 Detexmination of the minimum inhibitinq concentration 62 g. of a dispersion of a co-polymer of vinyl acetate and vinyl propionate, 16 g. polyvinylpyrrolidone dissolved in 22 ml. water, 1 ml. of the phosphate buffer described in Example 1 and 1 g. polyethylene sorbitan mono-oleate are stirred until homogeneous and the pH value adjusted to 7.3 with aqueous sodium hydroxide solution.
Into this film mass are stirred the antibiotics according to German Industrial Standard No. 58940 in increasing amounts, namely, 2.0 ~g., 20 ~g., 200 ~g., 2.0 mg and 20 mg. per 100 g.
The crude masses thus produced are, as described in Example 2, coated with a layer thickness of 350~ on to a nylon mesh of about 6ql thickness and 40~ filament thick-ness and subsequently dried at 50C. Test strips are produced from the "antibiotic films" thus obtained. For this purpose, on to a stiff foil there is laid a 2.5 x 2.5 cm.
nutrient card with Ca-So medium. Over the nutrient card there i~ sealed the '`antibiotic films". There can be pro-duced test strips for each antibiotic and each concentration or on to one test strip there can be applied fllms with various concentrations of the same antibiotic.
If these test strips are dipped, for example, into micro-organism-containing urines or if micro-organism sus-pensions are applied to the test zones, followed by incubation in known manner, the minimum inhibiting con-centration can be detsrmined without difficulty and from this the resistance of the micro-organism can be assessed and the correct therapeutic procedure instituted.
llZ l~ ~33 7 Determinations o~ the minimum inhibiting concent-ration carried out parallel thereto according to the well-known process using nutrient broth with subsequent nephelo-metric evaluation showed that the same therapeutic con-sequences are provided by the results of the process accord-ing to the present invention and from the well-known process.
Example 7 Determination of-the minimum inhibitin~ concentration of antibiotics which are more stable in another ~H range that in the pH ran~e of optimum micro-oraanism qrowth.
a) Double film based on a dispersion.
It is known that, for example, cephazolin is stable in a pH range of from 5.2 to 5.6 for a comparatively longer period of time than in the pH range of from 7.0 to 7.5, i.e.
the pH range for the optimum growth of pathogenic micro-organisms in urine. In this case, double films are produced in the following manner:
On to an antibiotic-free film according to Example 2 there is coated, in a layer thickness of 35 ~, a crude film mass according to Example 6 w~ich has been adjusted with aqueous sodium hydroxide solution to a pH value of 5.2 to 5.6 and into which has been stirred increasing amounts of cephazolin, the following amount~ of antibiotic per 100 g.
crude film mass thereby being stirred in:
20 ~g., 200 ~g,, 2.0 mg., 20 mg. and 200 mg.
The further working up, drying andproduction of the test strips, as wel~ as the testing of the resistance behaviour of the micro-organisms in urine and of suspensions of micro-organisms is carried out in the manner described in Example 6.
~12~)~3~7 Determinationi~ of the minimum inhibiting concen-tration carried out parallel thereto according to the well-known processes gave the same results, as described in Example 6.
hDouble-layer films" produced in this manner showed, even after 12 months, more than 95% of the activity of the cephazolin contained therein, whereas a film in which the cephazolin was present at a pH of 7.0 had already lost over 5~/0 of its activity.
b) Double film from film formers which are not dispersions.
The second, antibiotic-containing film can, instead of being produced from a dispersion, also be made from a film former which liberates the antibiotic during the incubation phase but which is not a dispersion.
20 g. Homopolymeric ethylene oxide are dissolved in sufficient water to give 100 ml. To this iis added 1 g.
polyoxyethylene sorbitan oleate. The solution is adjusted with hydrochloric acid to the necessary pH value of 5.2 to 5.6, the amounts of cephazolin mentioned in Example 7 a) are added thereto and the mixture is stirred until homogeneous.
Further working up and the results obtained with such tests are the isame as those described in Example 7 a).
Exa ple 3 Test strips in which, in addition to macromolecular openers; nutrient materials are present in the film.
62 g. of a dispersion of a co-polymer of vinyl acetate and vinyl propionate are mixed with 37 g. of a solution composed of 37 g. polyvinylpyrrolidone, 13.0 g.
disodium hydrogen phosphate and 2.3 g, potassium dihydrogen phosphate in 500 ml, water into which are stirred 2.0 g lactose and 1.0 g, polyoxyethylene sorbitan monooleate, )837 Films are produced from the crude film in the manner described in Example 2 and, as there described, test strips are produced by combination with a nutrient card which is impregnated with a universal medium.
On such test ~trips, pathogenic micro-organisms of the urine show, in some cases, much better formed colonies than on the well-known agar nutrient media.
Example 9 Test strlps in which onlY nutrients are added to the film as openers.
A crude film mass is prepared with the following composition. into 100 g. of a dispersion of a co-polymer of vinyl acetate and vinyl propionate are stirred 10 ml. of 0.7 M phosphate buffer in which are dissolved 1.5 g. glucose and 1.5 g. lactose. Thereafter, 1.0 g. polyoxyethylene sorbitan monooleate are stirred in and the crude film mass s adjusted to a pH of 7.3 with aqueous sodium hydroxide solution.
Test strips are produced with this crude film mass in the manner described in Example 8.
~;; Of the pathogenic micro-organisms of urine, Pseudomonas aerugino9a grows on these test strips consider-ably better than on most others so that, with this test, it is possible to carry out an orienting examination for this micro-organism.
_ 26 -~12V837 By forming the film from an aqueous dispersion of the polymer, micro-fine channels containing the opener are formed between the originally dispersed resin particles;
and the resin particles melt or coalesce together over a sufficient area to provide a film having the necessary strength.
Exam~le 3 Determination and differentiation of micro-orqanisms Films produced in the manner described in Example 1 are cut up into squares with 5 cm. long sides.
~utrient cards of the same size are laid in a Petri dish and moistened with sterile water. As nutrient media, there can be used all nutrient and nutrient-inhibitor compositions known in microbiology, For the determinatlon of the micro-organism count, it is best to use a universal medium, for example, the Ca-So medium. For micro-organism differentiation, there is used an 1~2V83'7 elective or selective medium, for example MacConkey's medium for the substantial inhibition of the gram~positive flora, Slanetz and Bartley's enterococcal-selective medium, Pseudo-monas elective medium, Wilson and Blair's Salmonella selective medium and the like~
On to the moistened nutrient cards are laid films, the wetting agents of which are chosen in such a manner that they are compatible with the nutrient medium and a precisely pipetted amount of micro-organism suspension distributed thereon with a spatula.
After an incubation period of 12 to 48 hours at 30 to 37C., by counting the colonies, the micro-organism count can be determined and, by the growth behaviour of the micro-organisms on the various media, the type thereof can be identified.
Example 4 Diaqnosis of urinarY infections - For the diagnosis of a urinary infection, use is made of a test strip according to Example 1. The wetting agent of the film i9, as in Example 3, coordinated with the nutrient medium. For this purpose, a nutrient card is cut with the dimensions of 2.5 x 2.5 cm., laid on a 2.5 x 10 cm.
~ealing foil and the film sealed with a hot-seal roller in such a manner that it lies close to the nutrient card. It is preferable to employ one nutrient card with CLED medium and one with MacConkey medium, which are placed side by side.
When such a test strip is dipped into a freshly excreted middle stream of urine and then incubated, there are found for all pathogenic micro-org~nisms in the urine, especially Colibacteria, Enterococci, Proteus mirabilis, Proteus vulqaris, Proteus morqanii, Proteus rettqeri, Pseudomonas aeruqinosa, Candida albicans, Klebsiella, Aerobacter aeroqenes, 11.ZC~337 ~ ilococcus aureus and Citrobacter, the same results as -with the well-known nutrient agar. The great advantage of the new test system in comparison with agar is that the test strips are packed in a sterile manner, can be stored for year~ and only require a fraction of the storage space needed for agar tests and, in addition, destruction thereof by incineration is less laborious.
Example 5 Preparation of antibioqram~
In order to test the resistance behaviour of micro-organisms cultured according to Example 4, the following procedure is used:
The usual procedure i9 employed in which colonies are inoculated with a loop, slurried in sterîle physiological sodium chloride solution and 0.1 ml. thereof further worked up.
For this purpose, a round nutrient card, for example, contain-ing Muller-Hinton's medium, is moistened with sterila water, covered with a film of the same size, the suspension of micro-organism spread thereon with a spatula and an antibiotic test leaflet according to German Industrial Standard ~o.
58940 laid thereon. In contradistinction to nutrient agar, it is not necessary to press on the test leaflet since this itself adheres to the moist surface of the test strip. After in¢ubation, for all pathogenic micro-organisms of the urine there are observed, in the case of all antibiotics used, a resistance behaviour which is identical with incubation in normal agar nutrient substrates.
- 2~ -112~83~
Example_6 Detexmination of the minimum inhibitinq concentration 62 g. of a dispersion of a co-polymer of vinyl acetate and vinyl propionate, 16 g. polyvinylpyrrolidone dissolved in 22 ml. water, 1 ml. of the phosphate buffer described in Example 1 and 1 g. polyethylene sorbitan mono-oleate are stirred until homogeneous and the pH value adjusted to 7.3 with aqueous sodium hydroxide solution.
Into this film mass are stirred the antibiotics according to German Industrial Standard No. 58940 in increasing amounts, namely, 2.0 ~g., 20 ~g., 200 ~g., 2.0 mg and 20 mg. per 100 g.
The crude masses thus produced are, as described in Example 2, coated with a layer thickness of 350~ on to a nylon mesh of about 6ql thickness and 40~ filament thick-ness and subsequently dried at 50C. Test strips are produced from the "antibiotic films" thus obtained. For this purpose, on to a stiff foil there is laid a 2.5 x 2.5 cm.
nutrient card with Ca-So medium. Over the nutrient card there i~ sealed the '`antibiotic films". There can be pro-duced test strips for each antibiotic and each concentration or on to one test strip there can be applied fllms with various concentrations of the same antibiotic.
If these test strips are dipped, for example, into micro-organism-containing urines or if micro-organism sus-pensions are applied to the test zones, followed by incubation in known manner, the minimum inhibiting con-centration can be detsrmined without difficulty and from this the resistance of the micro-organism can be assessed and the correct therapeutic procedure instituted.
llZ l~ ~33 7 Determinations o~ the minimum inhibiting concent-ration carried out parallel thereto according to the well-known process using nutrient broth with subsequent nephelo-metric evaluation showed that the same therapeutic con-sequences are provided by the results of the process accord-ing to the present invention and from the well-known process.
Example 7 Determination of-the minimum inhibitin~ concentration of antibiotics which are more stable in another ~H range that in the pH ran~e of optimum micro-oraanism qrowth.
a) Double film based on a dispersion.
It is known that, for example, cephazolin is stable in a pH range of from 5.2 to 5.6 for a comparatively longer period of time than in the pH range of from 7.0 to 7.5, i.e.
the pH range for the optimum growth of pathogenic micro-organisms in urine. In this case, double films are produced in the following manner:
On to an antibiotic-free film according to Example 2 there is coated, in a layer thickness of 35 ~, a crude film mass according to Example 6 w~ich has been adjusted with aqueous sodium hydroxide solution to a pH value of 5.2 to 5.6 and into which has been stirred increasing amounts of cephazolin, the following amount~ of antibiotic per 100 g.
crude film mass thereby being stirred in:
20 ~g., 200 ~g,, 2.0 mg., 20 mg. and 200 mg.
The further working up, drying andproduction of the test strips, as wel~ as the testing of the resistance behaviour of the micro-organisms in urine and of suspensions of micro-organisms is carried out in the manner described in Example 6.
~12~)~3~7 Determinationi~ of the minimum inhibiting concen-tration carried out parallel thereto according to the well-known processes gave the same results, as described in Example 6.
hDouble-layer films" produced in this manner showed, even after 12 months, more than 95% of the activity of the cephazolin contained therein, whereas a film in which the cephazolin was present at a pH of 7.0 had already lost over 5~/0 of its activity.
b) Double film from film formers which are not dispersions.
The second, antibiotic-containing film can, instead of being produced from a dispersion, also be made from a film former which liberates the antibiotic during the incubation phase but which is not a dispersion.
20 g. Homopolymeric ethylene oxide are dissolved in sufficient water to give 100 ml. To this iis added 1 g.
polyoxyethylene sorbitan oleate. The solution is adjusted with hydrochloric acid to the necessary pH value of 5.2 to 5.6, the amounts of cephazolin mentioned in Example 7 a) are added thereto and the mixture is stirred until homogeneous.
Further working up and the results obtained with such tests are the isame as those described in Example 7 a).
Exa ple 3 Test strips in which, in addition to macromolecular openers; nutrient materials are present in the film.
62 g. of a dispersion of a co-polymer of vinyl acetate and vinyl propionate are mixed with 37 g. of a solution composed of 37 g. polyvinylpyrrolidone, 13.0 g.
disodium hydrogen phosphate and 2.3 g, potassium dihydrogen phosphate in 500 ml, water into which are stirred 2.0 g lactose and 1.0 g, polyoxyethylene sorbitan monooleate, )837 Films are produced from the crude film in the manner described in Example 2 and, as there described, test strips are produced by combination with a nutrient card which is impregnated with a universal medium.
On such test ~trips, pathogenic micro-organisms of the urine show, in some cases, much better formed colonies than on the well-known agar nutrient media.
Example 9 Test strlps in which onlY nutrients are added to the film as openers.
A crude film mass is prepared with the following composition. into 100 g. of a dispersion of a co-polymer of vinyl acetate and vinyl propionate are stirred 10 ml. of 0.7 M phosphate buffer in which are dissolved 1.5 g. glucose and 1.5 g. lactose. Thereafter, 1.0 g. polyoxyethylene sorbitan monooleate are stirred in and the crude film mass s adjusted to a pH of 7.3 with aqueous sodium hydroxide solution.
Test strips are produced with this crude film mass in the manner described in Example 8.
~;; Of the pathogenic micro-organisms of urine, Pseudomonas aerugino9a grows on these test strips consider-ably better than on most others so that, with this test, it is possible to carry out an orienting examination for this micro-organism.
_ 26 -~12V837 By forming the film from an aqueous dispersion of the polymer, micro-fine channels containing the opener are formed between the originally dispersed resin particles;
and the resin particles melt or coalesce together over a sufficient area to provide a film having the necessary strength.
Claims (30)
1. A device for microbiological purposes, comprising a nutrient card and a covering layer comprising a water-insoluble synthetic resin film modified with an opener, said opener being effective to render said film impermeable to micro-organisms but permeable to nutrients, said film having been produced from an aqueous dispersion of the synthetic resin.
2. A device according to claim 1, wherein the opener is a water-soluble or water-swellable compound.
3. A device according to claim 1, wherein the opener is selected from the group consisting of polyethylene glycol, polyethylene oxide, polyvinylpyrrolidone, polyvinyl alcohol, partially saponified polyvinyl ester, a co-polymer of vinyl-pyrrolidone and a vinyl ester and cellulose derivatives.
4. A device according to claim 1, wherein the opener is a hydroxyalkylcellulose.
5. A device according to claim 1, 2 or 3, wherein the synthetic resin film is selected from the group consisting of a homopolymer of vinyl acetate, a homopolymer of an acrylic acid ester, a co-polymer of vinyl propionate and vinyl acetate, vinyl propionate and vinyl chloride, vinyl acetate and a maleic acid ester, an acrylic acid ester with acrylo-nitrile and vinyl propionate and butadiene and styrene.
6. A device according to claim 1, wherein the synthetic resin film has a thickness of from 10 to 1000µ.
7. A device according to claim 1, 3 or 6, wherein the weight ratio of opener to synthetic resin is from 5:1 to 1:50.
8. A device according to claim 1, wherein the film contains a synthetic resin mesh.
9. A device according to claim 8, wherein the synthetic resin mesh has a thickness of from 10 to 200µ.
10. A device according to claim 8 or 9, wherein the synthetic resin mesh is provided with coloured markings at definite distances apart.
11. A device according to claim 1, 6 or 9, wherein the covering layer contains at least one of a wetting agent, buffer, salt and nutrient.
12. A device according to claim 1, 6 or 9, wherein the covering layer contains at least one of a wetting agent, buffer, salt and nutrient in an amount less than 15% of the weight thereof.
13. A device according to claim 1, 6 or 9, wherein the covering layer contains at least one substance that inhibits the growth of particular micro-organisms.
14. A device according to claim 1, 6 or 9, wherein the covering layer contains at least one substance that promotes the growth of particular micro-organisms.
15. A device according to claim 1, 6 or 9, wherein the covering layer contains an antibiotic.
16. A device according to claim 1, 6 or 9, wherein the nutrient card is a filter paper impregnated with a nutrient solution.
17. A device according to claim 1, 6 or 9, wherein the nutrient card contains at least one material selected from the group consisting of a wetting agent, a moisture-retaining agent, a swelling agent, a buffer, a salt and an antibiotic.
18. A device according to claim 1, wherein the nutrient card is disposed on a water-resistant substrate.
19. A device according to claim 18, wherein the nutrient card is held in a pocket formed by the substrate and the covering layer.
20. A device according to claim 19, wherein the pocket is formed by adhering or sealing projecting edges of the covering layer adjacent to the nutrient card on to the substrate.
21. A device according to claim 20, wherein the sub-strate is provided with an elongation which serves as a handle.
22. A device according to claim 18, 20 or 21, wherein the substrate comprises a synthetic resin foil.
23. A device according to claim 18, wherein a plurality of covering layers and nutrient cards are disposed on the sub-strate to provide spaced apart test zones of different composition.
24. A device according to claim 23, wherein the com-positions differ in the concentration of an antibiotic in at least one of said covering layer and said nutrient card.
25. A test device for microbiological investigations comprising a water resistant substrate, a covering layer having peripheral edges secured to said substrate to define a pocket, and a nutrient card firmly held in said pocket, said nutrient card comprising a filter paper impregnated with dry nutrients for bacterial systems, said covering layer comprising a water-insoluble synthetic resin film having a thickness of 10 to 1000µ modified with a water-soluble or water-swellable opener, which does not have a negative effect on bacterial growth, said opener being effective to render said film impermeable to bacteria but permeable to nutrients and substances which inhibit or suppress the growth of micro-organisms, said covering layer having a smooth outer surface adapted to firmly hold bacteria in relative amounts correspond-ing to their concentrations on a medium to be investigated;
the weight ratio of said synthetic resin to said opener being from 50:1 to 1:5 and said film being derived from an aqueous dispersion of the synthetic resin.
the weight ratio of said synthetic resin to said opener being from 50:1 to 1:5 and said film being derived from an aqueous dispersion of the synthetic resin.
26. A test device according to claim 25, sealed between protective packing layers for storage.
27. A test device according to claim 25 or 26, wherein the film contains a synthetic resin mesh having a thickness of 10 to 200µ.
28. A method for determining micro-organism counts and for the differentiation of micro-organisms, wherein a device as defined in claim 1, 6 or 9, is contacted with a micro-organism-containing liquid to be investigated, excess liquid is removed, the device is incubated and the colonies of micro-organisms formed are counted or the types of micro-organisms are differentiated by the growth behaviour thereof on various media.
29. A method of producing an antibiogram, wherein a device according to claim 1, 6 or 9, is moistened with a suspension of micro-organisms, antibiotic test leaflets are laid thereon, followed by incubation, after which the resistance behaviour is determined.
30. A method for the determination of the minimum inhibiting concentration of antibiotics, wherein a device according to claim 1, 8 or 9, in which at least one of the covering layer and the nutrient card contains definite con-centrations of an antibiotic, is contacted with a micro-organism containing liquid, whereafter the device is incubated and the minimum inhibiting concentration of the antibiotic is determined.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE19782825636 DE2825636A1 (en) | 1978-06-12 | 1978-06-12 | DEVICE FOR MICROBIOLOGICAL WORK |
| DEP2825636.2 | 1978-06-12 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1120837A true CA1120837A (en) | 1982-03-30 |
Family
ID=6041568
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA000328718A Expired CA1120837A (en) | 1978-06-12 | 1979-05-30 | Device for microbiological purposes |
Country Status (15)
| Country | Link |
|---|---|
| US (1) | US4250256A (en) |
| EP (1) | EP0006192B1 (en) |
| JP (1) | JPS54163882A (en) |
| AR (1) | AR219607A1 (en) |
| AT (1) | ATE19T1 (en) |
| AU (1) | AU4778479A (en) |
| BR (1) | BR7903692A (en) |
| CA (1) | CA1120837A (en) |
| DD (1) | DD144277A5 (en) |
| DE (2) | DE2825636A1 (en) |
| DK (1) | DK242579A (en) |
| FI (1) | FI63059C (en) |
| PL (1) | PL216220A1 (en) |
| PT (1) | PT69753A (en) |
| ZA (1) | ZA792823B (en) |
Families Citing this family (33)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE2910134A1 (en) * | 1979-03-15 | 1980-09-25 | Boehringer Mannheim Gmbh | DIAGNOSTIC AGENT FOR DETECTING COMPONENTS OF BODY LIQUIDS |
| US4565783A (en) * | 1981-01-27 | 1986-01-21 | Minnesota Mining And Manufacturing Company | Dry culture media |
| JPH0249705B2 (en) * | 1981-01-27 | 1990-10-31 | Minnesota Mining & Mfg | |
| DE3136695A1 (en) * | 1981-09-16 | 1983-06-09 | Boehringer Mannheim Gmbh, 6800 Mannheim | DEVICE FOR DETECTING MICROORGANISM INHIBITING SUBSTANCES |
| JPS5863382A (en) * | 1981-10-13 | 1983-04-15 | Terumo Corp | Multi-layer culture test tool for microorganism |
| JPS58179499A (en) * | 1982-04-13 | 1983-10-20 | Wako Pure Chem Ind Ltd | Stabilization of disc for bacterial sensitivity test |
| DE3247608A1 (en) * | 1982-12-23 | 1984-07-05 | Boehringer Mannheim Gmbh, 6800 Mannheim | TEST STRIP |
| US4532107A (en) * | 1984-03-26 | 1985-07-30 | Miles Laboratories, Inc. | Reagent test device |
| US4717667A (en) * | 1984-07-11 | 1988-01-05 | Fmc Corporation | Colony replicating device |
| JPS6178397A (en) * | 1984-09-22 | 1986-04-21 | Kao Corp | Reaction process of enzyme and microorganism |
| US4656130A (en) * | 1985-03-14 | 1987-04-07 | Yissum Research Development Company | Collagen coated cell growth plates |
| SE456246B (en) * | 1985-08-01 | 1988-09-19 | Biodisk Ab | DEVICE FOR SENSITIVITY DETERMINATION OF MICRO-ORGANISMS INCLUDING A NON-POROS TEST STRAP WITH TWO DIFFERENT TESTS |
| DE3616496A1 (en) * | 1986-05-16 | 1987-11-19 | Boehringer Mannheim Gmbh | ANALYZING ELEMENT FOR DETERMINING COAGLING PARAMETERS |
| US4851210A (en) * | 1986-05-22 | 1989-07-25 | Genelabs Incorporated | Blood typing device |
| DE3640114A1 (en) * | 1986-11-24 | 1988-06-01 | Volker Barkey | Device for demonstrating the efficacy of a disinfection process when disinfectants are used in a washing process |
| US5240601A (en) * | 1988-11-09 | 1993-08-31 | Chembiomed, Ltd. | Affinity supports for hemoperfusion |
| US5149425A (en) * | 1988-11-09 | 1992-09-22 | Chembiomed, Ltd. | Affinity supports for hemoperfusion |
| JPH02167067A (en) * | 1988-12-21 | 1990-06-27 | Kirin Brewery Co Ltd | Sheet-like culture medium |
| US5089413A (en) * | 1989-05-19 | 1992-02-18 | Minnesota Mining And Manufacturing Company | Method and apparatus for culturing with microbiological dry culture medium |
| MY136048A (en) * | 1992-05-22 | 2008-08-29 | Chong Sue Kheng | Culturing of micro-organisms |
| DE69329424T2 (en) * | 1992-11-06 | 2001-04-19 | Biolog Inc | TEST DEVICE FOR LIQUID AND SUSPENSION SAMPLES |
| TW369562B (en) | 1996-07-17 | 1999-09-11 | Chisso Corp | Devices for detecting microorganisms and culture media for use in said devices |
| US6325980B1 (en) * | 1998-05-07 | 2001-12-04 | Serim Research Corporation | Test strip incubation device |
| WO2000023792A1 (en) * | 1998-10-19 | 2000-04-27 | Cbd Technologies Ltd. | Methods of concentrating microorganisms using affinity separation |
| US6583722B2 (en) | 2000-12-12 | 2003-06-24 | Kimberly-Clark Worldwide, Inc. | Wetness signaling device |
| US6603403B2 (en) | 2000-12-12 | 2003-08-05 | Kimberly-Clark Worldwide, Inc. | Remote, wetness signaling system |
| US20050081793A1 (en) * | 2003-10-16 | 2005-04-21 | Sannikka Martti J. | Container for urine |
| CN101076601B (en) | 2004-12-13 | 2013-11-13 | 拜尔保健有限公司 | Size self-limiting compositions and test devices for measuring analytes in biological fluids |
| AR057272A1 (en) * | 2005-04-04 | 2007-11-28 | Du Pont | FLEXIBLE CULTURE HANDBAG CONTAINING A NUTRIENT CONCENTRATE |
| US9939431B2 (en) * | 2013-03-29 | 2018-04-10 | Nima Labs, Inc. | Portable device for detection of harmful substances |
| US10249035B2 (en) | 2013-03-29 | 2019-04-02 | Nima Labs, Inc. | System and method for detecting target substances |
| US10254279B2 (en) | 2013-03-29 | 2019-04-09 | Nima Labs, Inc. | System and method for detection of target substances |
| US10466236B2 (en) | 2013-03-29 | 2019-11-05 | Nima Labs, Inc. | System and method for detecting target substances |
Family Cites Families (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US2672431A (en) * | 1949-11-25 | 1954-03-16 | Goetz Alexander | Means for performing microbiological assays of aerosols and hydrosols |
| US2672432A (en) * | 1951-03-23 | 1954-03-16 | Goetz Alexander | Means for performing microbiological assays of aerosols and hydrosols |
| US2904474A (en) * | 1954-10-04 | 1959-09-15 | Bacto Strip A G | Process and means for carrying out bacteriological operations |
| US3000706A (en) * | 1958-04-22 | 1961-09-19 | Boots Pure Drug Co Ltd | Control of bacteriological sterilisation |
| US3881993A (en) * | 1971-03-16 | 1975-05-06 | Miles Lab | Testing device for microorganisms |
| DE2118455B1 (en) * | 1971-04-16 | 1972-09-21 | Boehringer Mannheim Gmbh | Test strips |
| US3843452A (en) * | 1972-04-26 | 1974-10-22 | Miles Lab | Microbiological test article |
| US3814670A (en) * | 1972-04-26 | 1974-06-04 | Miles Lab | Test article for use in microbiology |
| FR2281982A1 (en) * | 1974-08-12 | 1976-03-12 | Api System | ANALYSIS DEVICE FOR THE RAPID SCREENING OF MICRO-ORGANISMS FROM AN ISOLATION ENVIRONMENT |
| FR2374353A1 (en) * | 1976-12-14 | 1978-07-13 | Comp Generale Electricite | POLYACRYLIC ACID MEMBRANES FOR ELECTROCHEMICAL USE |
-
1978
- 1978-06-12 DE DE19782825636 patent/DE2825636A1/en not_active Withdrawn
-
1979
- 1979-05-30 CA CA000328718A patent/CA1120837A/en not_active Expired
- 1979-06-01 US US06/044,629 patent/US4250256A/en not_active Expired - Lifetime
- 1979-06-05 AR AR276807A patent/AR219607A1/en active
- 1979-06-05 AU AU47784/79A patent/AU4778479A/en not_active Abandoned
- 1979-06-07 ZA ZA792823A patent/ZA792823B/en unknown
- 1979-06-07 AT AT79101806T patent/ATE19T1/en not_active IP Right Cessation
- 1979-06-07 EP EP79101806A patent/EP0006192B1/en not_active Expired
- 1979-06-07 JP JP7068579A patent/JPS54163882A/en active Pending
- 1979-06-07 DE DE7979101806T patent/DE2960171D1/en not_active Expired
- 1979-06-08 FI FI791846A patent/FI63059C/en not_active IP Right Cessation
- 1979-06-08 DD DD79213490A patent/DD144277A5/en unknown
- 1979-06-09 PL PL21622079A patent/PL216220A1/xx unknown
- 1979-06-11 BR BR7903692A patent/BR7903692A/en unknown
- 1979-06-11 PT PT69753A patent/PT69753A/en unknown
- 1979-06-11 DK DK242579A patent/DK242579A/en unknown
Also Published As
| Publication number | Publication date |
|---|---|
| EP0006192A1 (en) | 1980-01-09 |
| BR7903692A (en) | 1980-02-05 |
| ATE19T1 (en) | 1981-03-15 |
| FI63059C (en) | 1983-04-11 |
| PT69753A (en) | 1979-07-01 |
| DK242579A (en) | 1979-12-13 |
| PL216220A1 (en) | 1980-03-24 |
| DE2960171D1 (en) | 1981-04-02 |
| US4250256A (en) | 1981-02-10 |
| AU4778479A (en) | 1979-12-20 |
| JPS54163882A (en) | 1979-12-26 |
| AR219607A1 (en) | 1980-08-29 |
| DE2825636A1 (en) | 1979-12-20 |
| EP0006192B1 (en) | 1981-02-25 |
| ZA792823B (en) | 1980-07-30 |
| DD144277A5 (en) | 1980-10-08 |
| FI791846A (en) | 1979-12-13 |
| FI63059B (en) | 1982-12-31 |
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